Hepatitis C virus (HCV) is an important human pathogen that previously could be studied only in the chimpanzee model because the virus could not be grown in cultured cells. Growth of the virus in cultured hepatoma cells was recently achieved by Wakita et al. but even now growth is strain-specific and inefficient and infectious titers rarely reach even 100,000 viruses per ml of culture medium. Prior to discovery of this culture system, a retrovirus pseudotyped particle system was the only means to study neutralization of HCV and to perform mutational analysis of the two glycoproteins of HCV. In FY2008 we performed mutagenesis studies, utilizing pseudoparticles and infectious virus, that identified amino acid residues that are important for incorporation of the glycoproteins into infectious particles. For these studies, we developed a unique, single-cycle infection system that allowed us to identify mutations that specifically affected entry of HCV into cells. Entry into cells is controlled by the E1 and E2 glycoproteins of HCV. A number of neutralizing antibodies to the E2 glycoprotein have been identified in the past. We used both the pseudoparticle system and the cell culture system to demonstrate that the E1 glycoprotein is also a target of neutralizing antibodies and we showed that antibodies to a conserved region of E1 were broadly neutralizing.[unreadable] We extended our studies of enhancement of HCV infection by the human apolipoprotein C1 and demonstrated that this protein interacts with newly synthesized HCV within the host cell and that this association is maintained as viruses are released into cell culture medium or into the blood stream of infected animals. Thus, this host protein may influence both entry into and egress from host cells.